Project Summary/Abstract In spite of massive amounts of work, the neural basis of compulsive behavior in anxiety and especially addiction remains poorly understood. Much progress has been made in recent years in understanding the motivational role of drugs as positive incentives and rewards. Although it has long been known that aversive motivation also plays a role in addiction, this is less clearly understood. External stimuli associated with environmental stress or drug withdrawal are negative reinforcers that contribute to instrumental drug seeking and consumption responses by strengthening behaviors that allow escape from and/or avoidance of the aversive states elicited by these stimuli. Because active avoidance conditioning is based on negative reinforcement and involves brain circuits that overlap with addiction, we argue that a detailed understanding of the neural basis of escape/avoidance behavior will provide important information that may allow a deeper understanding of the role of aversive states in substance abuse. While much research was conducted on the neural basis on avoidance in the 1950s and 60s, this work fell out of favor, in part because the results did not lead to a clear understanding of the circuitry. However, in the intervening years, the neural basis of the first phase of avoidance, Pavlovian fear conditioning, has been elucidated in detail. This information makes it possible to revisit the neural basis of avoidance in a new light. In particular, given that we now understand in detail the neural mechanisms through which a neutral environmental stimulus associated with an aversive unconditioned stimulus (US) becomes a Pavlovian conditioned stimulus (CS) that elicits aversive states, we can now build on this information to understand the neural basis of avoidance conditioning. This is especially true if the same stimuli used as CSs and USs (tone and shock) are used to reveal the neural mechanisms of Pavlovian conditioning are also used in avoidance conditioning. The previously funded grant examined the contribution of the amygdala, a key structure for Pavlovian aversive conditioning, to avoidance. In this proposal we continue to pursue the role of the amygdala, but in addition also begin to dissect the broader circuitry involved. Specifically, we examine the role of connections between the subareas of the amygdala and nucleus accumbens, in the transition from Pavlovian conditioned reactions to negatively reinforced avoidant actions. Optogenetic techniques will be used to relate activity in specific amygdalostriatal pathways to discrete stages of avoidance learning and behavior, including precise negative reinforcement events (i.e. CS-termination, US- omission or both). Lastly, because dysfunction in nucleus accumbens endocannabinoid signaling may promote negative reinforcement and compulsions, we will use biochemistry, pharmacology and receptor knockdowns to examine the contribution that endocannabinoid signaling makes to negatively-reinforced avoidance responses.